The proposed research is planned to provide a comprehensive approach to the identification of cytoplasmic genes involved in cancer as well as in normal cellular processes in mammalian cells. Mitochondrial gene mutations will be sought by mutagenic treatments favoring mitochondrial mutations followed by suitable selection procedures. Putative cytoplasmic mutations will be tested by enucleation and cybrid formation or by reconstitution of cytoplasts from one cell line with karyoplasts from another, using pairs of cell lines carrying suitable genetic markers for selection. Similar procedures are being used to look for the cytoplasmic location of genes involved in tumorigenicity and its suppression. Cell lines being investigated include Balb/c 3T3 vs SVT2 (an SV40 transformed Balb/c mouse embryo fibroblast cell line); and virally transformed, chemically transformed, and spontaneously arising transformed cell lines arising from a non-transformed diploid Chinese hamster cell line (CHEF/18) developed in this laboratory. Non-mitochondrial cytoplasmic DNAs, seen in electron microscope studies of circular DNAs from mammalian cells, will be examined to lo k for sequence homology with either nuclear or mitochondrial DNAs.